Rotenone is a five-ring rotenoid compound found in the roots of several different plant species (Soloway (1976) Environ. Health Perspect. 14: 109-117). Likely used for centuries by indigenous peoples as a piscicide (fish poison) in the form of crude plant extracts and first purified more than a century ago, rotenone exhibits potent insecticidal, piscicidal, and pesticidal activities and is frequently used for these purposes (Ambrose & Haag (1936) Ind. Eng. Chem. Res. 28: 815-821; Geoffroy (1895) Ann. Inst. Colon. Marseille 2: 1-86; Soloway (1976) Environ. Health Perspect. 14: 109-117). Rotenone is listed as an active ingredient in more than forty commercially available pesticides approved for use by the U.S. Environmental Protection Agency.
It has been appreciated for some time that agricultural workers subjected to frequent pesticide exposure develop Parkinson's disease at a statistically higher rate than the general population, fueling suspicions that pesticides and other environmental toxins might contribute to the onset of this debilitating neurodegenerative disease, which afflicts as many as 1.5 million individuals in the United States alone (DeLong & Juncos (2005) In Harrison's Principles of Internal Medicine, D. Keal, ed. (New York, McGraw-Hill), pp. 2406-2418; Uversky (2004) Cell Tissue Res. 318: 225-241). It has also been shown that rats chronically treated with rotenone develop behavioral symptoms and pathophysiologies remarkably similar to those characteristic of human Parkinson's disease patients (Betarbet et al., (2000) Nat. Neurosci. 3: 1301-1306).
Rotenone is a specific inhibitor of complex 1 of the mitochondrial electron transport chain found in animal cells, which accounts for its pesticidal potency (Yagi et al., (1998) Biochim. Biophys. Acta 1364: 125-133). Dopaminergic neurons of rotenone-treated rats have been shown to exhibit mitochondrial abnormalities similar to those detected in the brains of deceased Parkinson's disease patients, including indications of complex I inhibition, and significant oxidative stress damage (Greenamyre et al., (2003) Parkinsonism Relat. Disord. 9 (Suppl 2): S59-64; Schapira, (2008) Lancet Neurol. 7: 97-109). While correlative evidence supports the notion that complex I inhibition might be at least partially responsible for dopaminergic cell death in idiopathic Parkinson's disease patients, evidence directly supportive of such a link has not been obtained. However, it has been shown that rotenone can induce dopaminergic neuron death independently of complex I, indicating the potential of complex I-independent mechanisms of rotenone-induced mitochondrial perturbation and cytotoxicity (Choi et al., (2008) Proc. Natl. Acad. Sci. U.S.A. 105:15136-15141).
Mitochondrial dysfunction is a primary or contributory factor in a broad range of neurological disorders and other human diseases, including Friedreich's ataxia, MALAS syndrome, and Barth syndrome, and is strongly implicated in the etiology of Parkinson's disease, a debilitating neurodegenerative disorder that afflicts millions of individuals worldwide. It is strongly suspected that most cases of sporadic Parkinson's disease, by far the most common form, arise from a combination of genetic predispositions and environmental factors, in particular exposures to pesticides and other environmental toxins. Supporting this hypothesis, it has been shown that prolonged treatment of rats with the widely used pesticide, rotenone, induces Parkinson's disease-like symptoms and pathophysiologies, including the mitochondrial dysfunction that is characteristic of the disease. While the rat-rotenone model now serves as an important tool for investigating the pathophysiology and potential treatment of Parkinson's disease, the system is not well suited for large-scale screens for identifying neuroprotective genes and drugs.
The fission yeast, Schizosaccharomyces pombe, and budding yeast, Saccharomyces cerevisiae, are species of fungi that do not possess the complex I components characteristic of animal cell mitochondria (Kerscher, (2000) Biochim. Biophys. Acta. 1459: 274-283). S. cerevisiae utilizes distinct NADH dehydrogenases, one exposed to the mitochondrial intermembrane space (NdeI) and the other to the mitochondrial matrix (NdiI), which allow for the oxidation of cytoplasmic and mitochondrial matrix NADH, respectively (Kerscher, (2000) Biochim. Biophys. Acta. 1459: 274-283). The S. pombe genome also encodes homologs NdeI and NdiI and it is presumed that they play roles analogous to their counterparts in budding yeast (Chiron, (2007) Methods Mol. Biol. 372: 91-105). Previous studies dating back to the early 1970s suggested that rotenone has little or no effect on mitochondrial function in either S. pombe or S. cerevisiae and it has since been largely accepted as dogma that both organisms have rotenone-resistant mitochondrial function (Chiron et al., (2007) Methods Mol. Biol. 372: 91-105; Kerscher, (2000) Biochim. Biophys. Acta. 1459: 274-283). While this dogma is supported by experimental evidence in the case of S. cerevisiae, the same cannot be claimed for S. pombe. The effects of rotenone on growth and mitochondrial function in this organism have been described only once (Heslot et al., (1970) J. Bacteriol. 104: 473-481), which showed that NADH dehydrogenase activity in this yeast is unaffected by relatively high concentrations of rotenone.